Hepatocellular carcinoma (HCC) is the most common malignant tumor of the liver and the third leading cause of death from cancer. However, currently there is no effective treatment for HCC. It is therefore urgent to develop novel therapeutic approaches for the treatment of HCC. Increasing evidence suggests that being able to modulate specific miRNAs may lead to the development of novel cancer therapies. Our studies (and others) indicate that a microRNA, miR-26a, represses proinflammatory cytokines, especially interleukin 6 (IL-6) and its downstream mediator, signal transducer and activator of transcription 3 (STAT3), suggesting a tumor suppressive role of miR-26a on HCC development. However, the role of miR-26a in HCC development in the setting consisting of both hepatocytes and Kupffer cells in appropriate HCC mouse models has not been determined. Furthermore, whether modulating miR-26a in hepatocytes and/or Kupffer cells can be an effective therapeutic approach for treating HCC has not been tested. The objective of this proposal is to investigate the molecular and cellular mechanisms by which miR-26a overexpression in hepatocytes and Kupffer cells inhibits HCC development. In addition, we have identified small molecules and developed siRNA/RNA in vivo targeted delivery technology that can increase miR-26a expression in the liver. In Aim 1, we will determine the effect of overexpression of miR-26a in hepatocytes and in Kupffer cells on suppression of HCC development. We have generated hepatocyte-specific as well as Kupffer cell-specific miR-26a overexpression transgenic mice. We will use these unique transgenic mouse lines to define the cellular and molecular mechanisms by which miR-26a exerts its effect on HCC. In Aim 2, we will determine the effect of small molecules on miR-26a induction and HCC suppression. We have also developed an innovative siRNA/RNA in vivo targeted delivery technology-- CpG-siRNA/RNA?that enables RNA in vivo delivery into Toll-like Receptor 9-positive Kupffer cells and inflammatory/malignant hepatocytes. CpG-Stat3 siRNA is moving towards clinical trials for glioma and lymphoma patients at City of Hope. We will assess the effect of both CpG-miR-26a and CpG-Stat3 siRNA on HCC development in hepatocytes or/and Kupffer cells in animal models. The proposed studies will not only provide mechanistic insights into pathways underlying HCC but also help to develop effective treatment strategies for HCC.